Prostate cancer is primarily fatal after it becomes metastatic and castration-resistant despite novel combined hormonal and chemotherapeutic regimens. Hence, new therapeutic concepts and drug delivery strategies are urgently needed for the eradication of this devastating disease. Here we report the highly specific, in situ click chemistry driven pretargeted delivery of cytotoxic drug carriers to PSMA(+) prostate cancer cells. Anti-PSMA 5D3 mAb and its F(ab')2 fragments were functionalized with trans-cyclooctene (TCO), labeled with a fluorophore, and used as pretargeting components. Human serum albumin (ALB) was loaded with the DM1 antitubulin agent, functionalized with PEGylated tetrazine (PEG4-Tz), labeled with a fluorophore, and used as the drug delivery component. The internalization kinetics of components and the therapeutic efficacy of the pretargeted click therapy were studied in PSMA(+) PC3-PIP and PSMA(-) PC3-Flu control cells. The F(ab')2 fragments were internalized faster than 5D3 mAb in PSMA(+) PC3-PIP cells. In the two-component pretargeted imaging study, both components were colocalized in a perinuclear location of the cytoplasm of PC3-PIP cells. Better colocalization was achieved when 5D3 mAb was used as the pretargeting component. Consecutively, the in vitro cell viability study shows a significantly higher therapeutic effect of click therapy in PC3-PIP cells when 5D3 mAb was used for pretargeting, compared to its F(ab')2 derivative. 5D3 mAb has a longer lifetime on the cell surface, when compared to its F(ab')2 analogue, enabling efficient cross-linking with the drug delivery component and increased efficacy. Pretargeting and drug delivery components were cross-linked via multiple bioorthogonal click chemistry reactions on the surface of PSMA(+) PC cells forming nanoclusters, which undergo fast cellular internalization and intracellular transport to perinuclear locations.

中文翻译：

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PSMA阳性前列腺癌中生物正交靶向治疗药物的细胞递送。

尽管有新型的激素和化学疗法联合治疗，前列腺癌在转移和去势抵抗后仍是致命的。因此，迫切需要新的治疗概念和药物递送策略来根除这种毁灭性疾病。在这里，我们报告高度特异性的原位点击化学驱动细胞毒性药物载体向PSMA（+）前列腺癌细胞的预先靶向递送。抗PSMA 5D3 mAb及其F（ab'）2片段用反式环辛烯（TCO）功能化，用荧光团标记，并用作预靶向组分。在人血清白蛋白（ALB）上加载DM1抗微管蛋白剂，并用PEG化四嗪（PEG4-Tz）功能化，并用荧光团标记，并用作药物递送成分。在PSMA（+）PC3-PIP和PSMA（-）PC3-Flu对照细胞中研究了组分的内在动力学和预靶向点击疗法的治疗功效。在PSMA（+）PC3-PIP细胞中，F（ab'）2片段的内在化速度比5D3 mAb快。在两成分预靶向成像研究中，两个成分都共定位在PC3-PIP细胞胞质的核周位置。当将5D3 mAb用作预靶向组分时，可以实现更好的共定位。连续地，体外细胞生存力研究显示，当使用5D3 mAb进行预靶向时，与其F（ab'）2衍生物相比，点击疗法在PC3-PIP细胞中具有显着更高的治疗效果。与F（ab'）2类似物相比，5D3 mAb在细胞表面的寿命更长，能够与药物输送成分有效地交联，并提高功效。预靶向和药物传递组件通过PSMA（+）PC细胞表面上的多个生物正交点击化学反应交联，形成纳米团簇，这些团簇经历快速的细胞内在化和细胞内转运至核周位置。